Background: Glioblastoma (GBM), the most common primary brain tumor, is the most aggressive malignancy in humans. Its rapid proliferation is a major obstacle to successful treatment. Patients with GBM often suffer from psychological disturbances associated with poor prognosis and physical discomfort. Diazepam is one of the most frequently used benzodiazepines (BZs) in cancer patients for its desirable psychotropic effects. The central effects of BZs are mediated by the activation of central BZ receptors. This study investigates whether diazepam has inhibitory effect on proliferation of GBM cell line T98G and explores its possible mechanism.
Methods: Cell viability and proliferation were respectively determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay and 5-bromo-2’-deoxyuridine (BrdU) incorporation assay. Cell cycle distribution was examined by flow cytometry. Western blot with specific protein antibodies was used to detect regulatory proteins involved in cell cycle regulation.
Results: Diazepam significantly decreased the proliferation of T98G cells in a dose-dependent and time-dependent manner. This effect was not reversed by the central BZ receptor antagonist flumazenil or the peripheral BZ receptor antagonist PK11195, indicating that it was not mediated by BZ receptors. Flow cytometry indicated that diazepam caused a cell accumulation in G0/G1 phase, thereby contributing to cell proliferation inhibition. Furthermore, our findings showed that lessened phosphorylation of Rb accounted for diazepam-induced G0/G1 phase arrest.
Conclusions: Diazepam inhibits the proliferation of human GBM T98G cells by inducing G0/G1 phase arrest. Diazepam has potential to be a lead for new drugs in GBM therapy because of its antitumor activity.
Departments of †Neonatology
‡Anesthesiology, Sun Yat-sen Memorial Hospital
Departments of *Pharmacology
∥Guangzhou Cellproteck Pharmaceutical Co. Ltd
¶Microbiology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, P.R. China
§Department of Anesthesiology, State University of New York-Downstate Medical Center, Brooklyn, NY
J.C. and Y.O. contributed equally. J.H., J.L., J.C., and Y.O. designed research; J.C., Y.O., L.C., Yuxi Zhou, Yuehan Zhou, X.Y., and H.Z. performed research; J.C., Y.O., L.C., W.Z., and G.Y. analyzed data; J.C. and Y.O. wrote the paper.
Supported by Key Program, National Natural Science Foundation of China (30830111), Guangzhou Program Project of Science Technology (2008Z1-E561), South China Comprehensive Platform for New Medicine R&D (2009ZX09301-015), and the Doctoral Fund of Ministry of Education of China (No. 20100171120052).
The authors have no conflicts of interest to disclose.
Reprints: Jun Hu, PhD, Department of Microbiology, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, 510080, P.R. China (e-mail: email@example.com) and Jun Lin, MD, PhD, Department of Anesthesiology, State University of New York - Downstate Medical Center, Brooklyn, NY 11203 (e-mail: firstname.lastname@example.org).
Received July 14, 2012
Accepted December 6, 2012